An ancient legacy of life transitioning from oceans to land is the role of the skeleton and kidney in regulating mineralization and Ca/PO4 homeostasis. Key players in this bone kidney axis are a group of related extracellular matrix proteins (SIBLINGs). These proteins all map to a tightly clustered region on chromosome 4q. A unique protein PHEX, regulates at least two SIBLING proteins (MEPE and DMP1), indirectly or directly. PHEX is a zn-metalloendopeptidase in search of a substrate and may also play a role as an orphan ligand or receptor. Loss of PHEX function results in X-linked hypophosphatemic rickets and a novel cytokine FGF23 (a phosphatonin), plays a major role in the phosphate phenotype. Our data confirms direct binding of PHEX to MEPE, a SIBLING protein expressed in bone, teeth and in the proximal convoluted tubules of the kidney. The binding between these proteins occurs between the PHEX Zn-binding motif and a short COOH-terminal region of MEPE called the ASARM-motif. This short region is exquisitely resistant to proteolysis and conserved across species. Biologically, the released ASARM-peptide is a potent inhibitor of mineralization, inhibits renal phosphate handling and binds to hydroxyapatite. Binding of PHEX to MEPE may serve to regulate release of the ASARM peptide and thus locally control mineralization. PHEX also binds with high affinity to free ASARM- peptide and also inactivates the peptide by hydrolysis. In HYP, as well as a loss of PHEX function, there is a marked up regulation of MEPE and osteoblastic protease activity. This results in degradation of MEPE and DMP1 and release of ASARM-peptides from both these proteins and perhaps other SIBLING proteins (DSPP, osteopontin BSP, enamelin). These protease resistant ASARM-peptides accumulate in kidney, bone, urine and in the circulation. To study the role of SIBLINGs, PHEX and ASARM- peptides we have made transgenic mice (MEPE-TRG) that overexpress MEPE protein 50X in bone and 7X in kidney. Our specific aims are: 1. Characterize bone and renal PO4 phenotypes of MEPE-TRG mice, 2. Determine the bone and renal- calcification roles of processed forms of MEPE and ASARM-peptides in MEPE-TRG, HYP and NPT2a-/- mice, 3. Design PHEX synthetic peptides as tools to probe the role of ASARM peptides and PHEX in health and disease.